Each of the references cited herein is incorporated by reference in its entirety. A complete listing of the citations is set forth at the end of the specification.
An estimated 211,300 new cases of invasive breast cancer are expected to occur among women in the United States during 2003. Breast cancer is the most frequently diagnosed non-skin cancer in women. (1) Estrogen is a steroid hormone that, while having important functions including the control of reproduction and the development of secondary sexual characteristics, also plays a predominant role in breast cancer growth and development. The use of estrogen for its positive effects can also detrimentally result in the stimulation of other tissues, such as those of the breast and uterus, and increase the risk of cancer at these sites.
The estrogen receptor (ER) is a member of a nuclear receptor superfamily consisting of orphan receptors and receptors for classic high-affinity ligands, such as steroid hormones, vitamin D, retinoids, and thyroid hormones. As a ligand inducible transcription factor, the estrogen receptor mediates the activity of estrogen in the development and function of the female reproductive system, the maintenance of bone mineral density, regulation of blood lipid profile, brain function, cardiovascular health and other physiologic processes.
Estrogen-Related Receptors (ERRs) are included in the nuclear receptor family and were the first orphan nuclear receptors found through a search for genes encoding proteins related to known nuclear receptors. While it was originally believed that the development and physiological roles of ERRs were quite distant from those of the classic ERs, it has recently been shown that in some cases ERRs can share target genes, coregulatory proteins, ligands, and sites of action with the ERs. (2) Like ER, ERRs are also implicated in breast cancer and other diseases. (3)
(Z)-Tamoxifen (Z-TAM), (Z)-2-[4-(1,2-diphenyl-1-butenyl) phenoxy]N,N-dimethylethanamine, is used clinically to treat estrogen-dependent breast cancer by acting as an antagonist of estrogen-induced tumor growth. The mechanism for its principal action is its competition with the natural agonist hormone estradiol (E2) for binding to the estrogen receptor ligand-binding domain, thereby reducing the ability of estradiol to stimulate nuclear transcription and consequent cell growth. For example, it is known that (Z)-4-hydroxytamoxifen (4-OHT), a potent tamoxifen metabolite, is a selective estrogen-receptor modulator that functions as an antagonist in breast cancer cells but displays estrogen-like activities in the uterus and bone. The Z-4-OHT form isomer has the required antiestrogenic activity, but E-4-OHT isomer has only about 5% of its affinity for ER. (17)
Selective Estrogen-Receptor Modulators (SERMs) are a type of estrogen receptor ligand that can exert agonist, antagonist, or neutral effects, depending on factors including the target gene and/or target tissue. (2) SERMs properties are related to their ability to compete in target tissues with estradiol for binding sites in the ligand-binding domain (LBD) of the ER. Tamoxifen and particularly its metabolite 4-hydroxytamoxifen are SERMs that also antagonize ERRs, but much higher doses of those SERMs are required to antagonize ERRs than ERs. In recent studies examining whether other ER ligands could influence ERR activity, (Z)-4-OHT was identified as the most potent isoform-specific inhibitor of ERRβ,γ. (4) The structures of (Z)-TAM, (Z)-4-OHT, and E2 are:

Recent experiments have synthesized 1,1-bis and 1,1,2-tris (4-hydroxyphenyl)-2-phenylalkene analogs for the studies of their antagonist action. (5) Studies also have shown that 1,1,2-triarylethenes are antagonists and that they inhibit the effect of 1 nN E2, dependent on the length of C2-alkyl chain. When C2 is substituted by an ethyl or trifluoromethyl group of the 1,1,2-triarylethene system, the substituted compound possesses the same antagonistic potency as 4-hydroxytamoxifen and is almost 50 times more active than tamoxifen itself. (6)
None of the 1,1-bis- and 1,1,2-tri (4-hydroxyphenyl)-2-phenylalkylenes bear a basic side chain, which is responsible for antagonistic effects of ER. Removal of the dimethylaminoethoxy side chain of 4-hydroxytamoxifen did not decrease the antagonistic effects on the MCF-7-2a cell line. This finding indicated that, in the class of 1,1-bis- and 1,1,2-tris (4-hydroxyphenyl)-2-phenylalkenes, having a basic side chain is not a prerequisite for exhibiting high binding affinity and antagonistic effects on ER. The antiestrogenic properties comprising estrogen receptor binding depend only on the length of the C2-alkyl chain.
Other experiments have also revealed information about the various ERRs. Estrogen Related Receptor Alpha (ERRα or ERRa or ERRa-1 or ERRα-1) binds the ligand diethylstilbestrol (DES) with greater affinity than 4-hydroxytamoxifen as antagonist/inverse agonist. ERRα functions to control adiposity and energy metabolism. In experiments with ERRα knockout mice, the mice are lean. ERRα also regulates the medium-chain acyl-CoA dehydrogenase (MCAD) gene in conjunction with PGC-1α (peroxisome proliferator-activated receptor gamma coactivator 1), a key regulator of lipid and glucose homeostasis.
PGC-1α is a transcriptional co-activator that regulates numerous pathways controlling both metabolism and overall energy homeostasis. (15) ERRα binding to PGC-1α requires the AF2 domain of ERRα. PGC-1α induces the expression of ERRα, meaning that ERRα is upregulated in response to signals that induce PGC-1α, such as exposure to cold. Expression of PGC-1α led to the induction of ERRα at the RNA and protein level in SAOS2-GR(+) cells, as well as in HtTA-1, HepG2, and 293 cells.
It has been found that PGC-1α and ERRα have a similar pattern of expression in human tissues, with both being present predominantly in organs with high metabolic needs such as skeletal muscle, kidney, and heart. Physiological stimuli such as fasting coordinately induces PGC-1α and ERRα. ERRα can dramatically and specifically repress PGC-1α transcriptional activity. (7)
ERRα is known to be expressed in breast cancer cells and it inhibits breast cancer cell growth independent of the estrogen receptor. ERRα regulates aromatase and pS2 genes and is associated with unfavorable biomarkers in human breast cancer. ERRα is also expressed in osteoblasts and regulates osteopontin expression. A recent study demonstrates that the ERRα gene is a downstream target of ERα. (16)
Further, ERRα has been shown to modulate oxidative phosphorylation gene expression that is altered in diabetic muscle. (25-27) It was suggested that ERRα agonists might ameliorate insulin-resistance in individuals with type 2 diabetes mellitus, and therefore that its antagonists could play roles in the development of diabetes.
Estrogen Related Receptor Beta (ERRβ or ERRb) binds the ligand 4-hydroxytamoxifen with a greater affinity than DES as antagonist/inverse agonist. ERRβ controls trophoblast proliferation and placental function. In mice lacking ERRβ, trophoblast stem cell differentiation is impaired and the placenta fails to develop normally. (8) ERRβ is present early in the developing placenta in a subset of cells in extra-embryonic ectoderm destined to make up the chorion. (8,9) Thus, ERRβ is likely essential for reproduction. ERRβ synthesis is highly restricted in postnatal life, being detected at low levels in the liver, stomach, skeletal muscle, heart and kidney. (5)
Estrogen Related Receptor Gamma (ERRγ or ERRg) binds the ligand 4-hydroxytamoxifen with a greater affinity than DES as antagonist/inverse agonist. ERRγ is expressed in heart, skeletal muscle, kidney, and brain as well as in the developing nervous system. Human ERRγ transcripts can be detected at very high levels in fetal brain, and at lower levels in fetal kidney, lung and liver. (4) In adult tissues, ERRγ is widely expressed and can be detected in brain, lung, bone marrow, adrenal and thyroid glands, trachea and spinal cord. In the mouse, the gene encoding ERRγ is expressed in specific areas of the brain, in addition to the heart, kidney, muscle, spleen and testis. With respect to its role in breast cancer, ERRγ associates with favorable biomarkers in human breast cancer and may regulate MCAD.
ERRα, ERRβ, and ERRγ do not respond to natural estrogens. They do recognize, however, the estrogen response element and can modulate gene expression in the absence of exogenously added ligand. 4-OHT disrupts the interaction between ERRβ and its co-regulator proteins and ERRγ and its co-regulator proteins. 4-OHT also abolishes the constitutive transcriptional activity of these receptors in transient transfection assays. In contrast, 4-OHT has no effect on coregulator/ERRα interaction or its transcriptional activity, demonstrating the existence of a novel nuclear receptor-based pharmacological pathway that may contribute to the tissue-specific activities of 4-OHT. (10)
In addition, we have found that found that ERRs can up-regulate aromatase expression in breast cancer cells. Therefore, antagonists of ERRs can be useful to treat breast cancer by the suppression of ERR activity and the reduction of estrogen formation. (21)
Thus, ERs and ERRs are targets of modulation. There is a pressing need, however, for compounds that modulate these receptors more effectively. There is also a need for methods of using these compounds to treat estrogen-related disorders.